Next-generation-based targeted sequencing in a set of Spanish Hirschsprung patients as an efficient tool for mutation detection.

The development of next-generation sequencing (NGS) technologies has a great impact in human mutation detection procedures given their high throughput nature. The purpose of this study was the validation of a designed panel of Hirschsprung disease (HSCR) associated genes as a rapid and efficient tool to perform a mutational screening in a series of patients. We have performed NGS-based targeted sequencing using a panel containing 26 associated or candidate genes for HSCR in a group of 11 selected Spanish HSCR patients carrying known mutations in any of the genes included in the panel. We have used the 454-GS Junior DNA sequencer as a rapid sample processing platform. The average percentage of covered bases was of 97% and 91.40% of the targeted bases were covered with depth = 20X. The mean coverage was 422X. In addition, we have found a total of 13 new coding variants and 11 new variants within regulatory regions among our patients. These outcomes let us deepen our knowledge of the different molecular events involved in the genetic basis of the disease in those patients. Here we report our validated NGS panel as an optimum method for the identification of new variants in our patients. This approach could be used as a research tool for a fast, reliable and safe mutational screening in another series of patients, even including newly associated genes linked to disease.